Abstract
X-linked inhibitor of apoptosis (XIAP) is a protein that possesses anti-apoptotic function and dysregulation of it has been linked to a number human disease such as cancers and neurodegenerative disorders. In our previous study, we have found that nitric oxide (NO) can modulate the anti-apoptotic function of XIAP and found that this can contribute to the pathogenesis of Parkinson’s disease. Specifically, we found that modification of baculoviral IAP repeat 2 of XIAP by S-nitrosylation can compromise XIAP’s anti-caspase 3 and anti-apoptotic function. In this study, we report that cysteine (Cys) 90, Cys 213 and Cys 327 can be specifically S-nitrosylated by NO. We found that mutations of Cys 90 and Cys 327 affect the normal structure of XIAP. More importantly, we found that S-nitrosylation of XIAP Cys 213 impairs the anti-caspase 3 and anti-apoptotic function of XIAP that we observed in our previous study.
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Acknowledgments
This work was supported by Hong Kong Research Grants Council Theme-based Research Scheme (T13-607/12R), HKUST10/CRF/12R, DAG12SC03S and HKUST12/CRF/13G.
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10495_2015_1087_MOESM1_ESM.eps
Supplementary material 1 Figure S1. Mutation of Cys90 of XIAP does not affect its E3 ligase activity but reduce its solubility. (A) HEK293T cells transfected with myc-tagged C90H XIAP and HA-Ub were incubated with or without 100 μM NOC18 for 24 h. The cells were then harvested and anti-myc antibody immunoprecipitation was performed on the soluble fraction followed by Western blot analysis to assess the E3 ligase activity of C90H XIAP. (B) Insoluble fraction of (A) was solubilized as described in the method section. The solubility of C90H XIAP was reduced as in Figure 1 B. (C) HEK293T cells transfected with myc-tagged C327H XIAP and HA-Ub were incubated with or without 100 μM NOC18 for 24 h. The cells were then harvested and anti-myc antibody immunoprecipitation was performed on the soluble fraction followed by Western blot analysis to assess the E3 ligase activity of C327H XIAP. No myc-tagged C327H XIAP was detected after immunoprecipitation. (D) Insoluble fraction of (C) was solubilized as described in the method section. The solubility of C327H XIAP was greatly reduced as in Figure 1 A (EPS 5647 kb)
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Wu, W., Wan, O.W. & Chung, K.K.K. S-nitrosylation of XIAP at Cys 213 of BIR2 domain impairs XIAP’s anti-caspase 3 activity and anti-apoptotic function. Apoptosis 20, 491–499 (2015). https://doi.org/10.1007/s10495-015-1087-3
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DOI: https://doi.org/10.1007/s10495-015-1087-3